Azithromycin analogue CSY0073 attenuates lung inflammation induced by LPS challenge.

BACKGROUND AND PURPOSE: Azithromycin is a macrolide antibiotic with anti-inflammatory and immunomodulating effects. Long-term azithromycin therapy in patients with chronic lung diseases such as cystic fibrosis has been associated with increased antimicrobial resistance, emergence of hypermutable strains, ototoxicity and cardiac toxicity. The aim of this study was to assess the anti-inflammatory effects of the non-antibiotic azithromycin derivative CSY0073. EXPERIMENTAL APPROACH: We compared the effects of CSY0073 with those of azithromycin in experiments on bacterial cultures, Pseudomonas aeruginosa biofilm, lung cells and mice challenged intranasally with P. aeruginosa LPS. KEY RESULTS: In contrast to azithromycin, CSY0073 did not inhibit the growth of P. aeruginosa, Staphylococcus aureus or Haemophilus influenzae and had no effect on an established P. aeruginosa biofilm. Bronchoalveolar lavage (BAL) fluids and lung homogenates collected after the LPS challenge in mice showed that CSY0073 and azithromycin (200 mg·kg(-1), i.p.) decreased neutrophil counts at 24 h and TNF-α, CXCL1 and CXCL2 levels in the BAL fluid after 3 h and IL-6, CXCL2 and IL-1ß levels in the lung after 3 h compared with the vehicle. However, only azithromycin reduced IL-1ß levels in the lung 24 h post LPS challenge. CSY0073 and azithromycin similarly diminished the production of pro-inflammatory cytokines by macrophages, but not lung epithelial cells, exposed to P. aeruginosa LPS. CONCLUSIONS AND IMPLICATIONS: Unlike azithromycin, CSY0073 had no antibacterial effects but it did have a similar anti-inflammatory profile to that of azithromycin. Hence, CSY0073 may have potential as a long-term treatment for patients with chronic lung diseases.

Cognitive procedural learning in early Alzheimer's disease: impaired processes and compensatory mechanisms.

INTRODUCTION: The aim of this study was to study cognitive procedural learning in early Alzheimer's disease (AD). METHODS: Cognitive procedural learning was assessed using the Tower of Hanoi (TH) task. In order to take account of possible interactions between different systems during cognitive procedural learning, we also measured non-verbal intellectual functions, working memory, and declarative memory. RESULTS: Our results showed an apparent preservation of cognitive procedural learning in AD and a deleterious effect of the disease on verbal intelligence and declarative memory. Correlational analyses revealed a difference between AD patients and control participants in the type of processing they applied to the task. CONCLUSION: The non-involvement of declarative memory would appear to be partly responsible for a slowdown in the cognitive procedural dynamics of AD patients. As the AD patients were unable to use their declarative memory, they were still in a problem-solving mode at the end of the learning protocol and had to implement higher order cognitive processes (i.e., compensatory mechanisms) to perform the procedural task.

Expression of miR-487b and miR-410 encoded by 14q32.31 locus is a prognostic marker in neuroblastoma.

BACKGROUND: Combination of age at diagnosis, stage and MYCN amplification stratifies neuroblastoma into low-risk and high-risk. We aimed to establish whether a microRNA (miRNA) signature could be associated with prognosis in both groups. METHODS: Microarray expression profiling of human miRNAs and quantitative reverse-transcriptase PCR of selected miRNAs were performed on a preliminary cohort of 13 patients. Results were validated on an independent cohort of 214 patients. The relationship between miRNA expression and the overall or disease-free survival was analysed on the total cohort of 227 patients using the log-rank test and the multivariable Cox proportional hazard model. RESULTS: A total of 15 of 17 miRNAs that discriminated high-risk from low-risk neuroblastoma belonged to the imprinted human 14q32.31 miRNA cluster and two, miR-487b and miR-410, were significantly downregulated in the high-risk group. Multivariable analyses showed miR-487b expression as associated with overall survival and disease-free survival in the whole cohort, independently of clinical covariates. Moreover, miR-487b and miR-410 expression was significantly associated with disease-free survival of the non-MYCN-amplified favourable neuroblastoma: localised (stage 1, 2 and 3) and stage 4 of infant <18 months. CONCLUSION: Expression of miR-487b and miR-410 shows predictive value beyond the classical high-/low-risk stratification and is a biomarker of relapse in favourable neuroblastoma.

[North African and Southeast Asian nasopharyngeal carcinomas: between the resemblance and the dissemblance]. / Le carcinome nasopharyngé en Afrique du Nord et dans le Sud-Est asiatique: entre similitude et différence.

Nasopharyngeal carcinoma (NPC) is an unusual head and neck cancer because of its unequal geographical distribution and its consistent association with the Epstein-Barr virus (EBV). This malignant tumor poses a serious public health problem in many countries, especially in Southeast Asia and North Africa where the recorded incidence are highest. During the past decade, a growing number of studies were undertaken to define the molecular basis of NPC. However, the analysis of several clinical and biological parameters of North African and Southeast Asian NPCs has shown notable differences, suggesting that they could result from a distinct combination of etiological factors. One intriguing characteristic of North African NPC, concerns its bimodal age distribution with a secondary peak of incidence in the range of 15-25 years, not observed in Asian NPC. In this juvenile form of NPC, immuno-histochemistry assay has shown that the two key proteins controlling the apoptotic-survival balance p53 and Bcl-2 are less frequently expressed whereas the transmembrane tyrosine-kinase receptor c-kit and the main EBV oncoprotein LMP1 were more abundant. In addition, the EBV serological alterations are less informative for the diagnosis of the juvenile compared to the adult form. In addition, most North African NPCs contain EBV strains with genetic polymorphisms distinct from those described in the Southeast Asia series (predominance of F, D, H1-H2, XhoI+ and f, C, H, XhoI- respectively). In contrast, studies relating on tumor chromosomal alterations or aberrant promoter methylation result in data very similar to those obtained from the Southeast Asia series, supporting the concept of a common molecular basis for all NPC regardless of patient geographic origin.

[Expression of c-kit in North African nasopharyngeal carcinomas: correlation with age and LMP1]. / Expression de c-kit dans les carcinomes nasopharyngés nord africains, corrélations avec l'âge et LMP1.

PURPOSE: To determine the level and prognostic significance of c-kit expression in the two age groups of North African nasopharyngeal carcinomas. PATIENTS AND METHODS: A retrospective study of 99 NPC specimens from Tunisian patients was investigated by immunohistochemistry. Immunohistochemical data were correlated with Epstein-Barr virus LMP1 expression and pathological, clinical and survival parameters. RESULTS: c-kit was detected in 79% of the cases for patients under 30 years of age (juvenile form) but in only 56% of specimens in patients over 30 years (P=0.039) and was significantly over-expressed for patients with lymph node involvement (P=0.015). LMP1 score was 5.78 (+/-1.84) for c-kit negative tumors compared to 8,23 (+/-2.39) for c-kit positive tumors (P=0.002). Multivariate analysis including age, lymph nodes involvement and LMP1 expression as co-variables, showed that only age (P=0.027) and LMP1 expression (P=0.005) were significantly correlated to the c-kit expression. CONCLUSION: c-kit is highly expressed in the juvenile form of North African nasopharyngeal carcinomas. There is a significant association between LMP1 and c-kit expression. The contrasted levels of C-kit expression in the two age groups strengthen the hypothesis that these clinical forms result from distinct oncogenic mechanisms.

[Nasopharyngeal carcinomas: from biology to clinic]. / Les carcinomes du nasopharynx: de la biologie à la clinique.

Nasopharyngeal carcinomas (NPC) are very different from other head and neck cancers because of their specific multifactorial etiology and their geographic distribution. Epstein-Barr Virus (EBV) is implicated in oncogenesis of NPC in association with genetic alterations such as inactivation of the p16/Ink4, p19/ARF, RASSF1 or Blu genes. Tumoral tissues include a very abundant characteristic lymphoid infiltrate. Inflammatory cytokines are produced by both malignant and infiltrating cells. There is no efficient immune response against the tumor. On the opposite, infiltrating lymphocytes might play a role in tumor development. Serological methods and detection of circulating viral DNA are expected to become useful for early detection of relapse and on a longer term for primary screening. NPC are often diagnosed at a late stage because patients may remain asymptomatic for a long time. Computed tomography (CT scan) and magnetic resonance imaging (MRI) are complementary for the initial evaluation. Positron emission tomography (PET) is efficient for the evaluation of treatment efficiency and detection of relapses. Treatment is based on radiotherapy and chemotherapy. Their optimal use needs to be evaluated by phase III trials but positive results have been obtained by concomitant association of radiotherapy and chemotherapy. Targeted therapies are being studied with strategies based on disruption of viral latency, use of replicative adenoviruses or anti-tumor vaccination.

Conventional and array-based comparative genomic hybridization analysis of nasopharyngeal carcinomas from the Mediterranean area.

Nasopharyngeal carcinoma (NPC) occurs with a high incidence in Southeast Asia and to a lesser extent in the Mediterranean area, especially in Tunisia, Algeria, and Morocco. Cellular gene alterations combined with latent Epstein-Barr virus infection are thought to be essential for NPC oncogenesis. To date, chromosome analysis with comparative genomic hybridization (CGH) has been reported exclusively for NPCs from Southeast Asia. Although NPCs from the Mediterranean area have several distinct clinical and epidemiological features, CGH investigations have been lacking. Chromosome analysis was therefore undertaken on a series of NPC xenografts and biopsies derived from patients of Mediterranean origin. Four xenografts were investigated with a combination of conventional CGH, array-based CGH, and comparative expressed sequence hybridization. In addition, 23 fresh NPC biopsies were analyzed with conventional CGH. Data obtained from xenografts and fresh biopsies were consistent, except that amplification of genes at 18p was observed only in xenografts derived from metastatic tissues. Frequent gains associated with gene overexpression were detected at 1q25 approximately qter (64%) and 12p13 (50%). Losses were noticed mainly at 11q14 approximately q23 (50%), 13q12 approximately q31 (50%), 14q24 approximately q31 (43%), and 3p13 approximately p23 (43%). Comparison with previous reports suggests that Mediterranean NPCs have higher frequencies of gains at 1q and losses at 13q than their Asian counterparts.

Nasopharyngeal carcinomas: an update.

Among the group of head and neck cancers, nasopharyngeal carcinomas (NPC) represent a distinct entity in terms of their epidemiology, clinical presentation, biological markers, carcinogenic risk factors, prognostic factors, treatment and outcome. Undifferentiated NPC (UCNT), the most frequent histological type, is endemic in certain regions, especially in South East Asia. The disease has also been associated with the presence of the Epstein-Barr Virus (EBV). Although NPC is a radiosensitive and chemosensitive tumour, a substantial number of patients develop local recurrence or distant metastases. For patients with locoregional advanced disease, it is well known that conventional radiotherapy is insufficient in terms of both the local control rates and distant metastases. New techniques of radiation and new combined radiotherapy and chemotherapy modalities have been evaluated in numerous clinical trials in recent years. The purpose of this article is to review the current knowledge in terms of the epidemiology, biology, prognosis, management and outcome of patients with NPC.

Adenovirus-p53 gene therapy in human nasopharyngeal carcinoma xenografts.

BACKGROUND: One major challenge to human cancer gene therapy, is efficient delivery of the gene-vector complex. METHODS AND RESULTS: Using two distinct human nasopharyngeal carcinoma (NPC) models, we demonstrate that intra-tumoural (IT) administration of adenoviral-mediated wild-type p53 gene therapy (Ad-p53) caused no greater inhibition of tumour growth as compared to ionizing radiation (XRT) alone. Detailed histologic examination of tumour sections demonstrated that <15% of tumour cells were transduced by IT adv-beta-gal. CONCLUSIONS: This report underscores the importance of developing gene transfer vectors, which can provide therapeutic levels of transgene expression efficiently in solid tumours.

Growth transformation of primary epithelial cells with a NPC-derived Epstein-Barr virus strain.

The Epstein-Barr virus (EBV) is associated with two major human epithelial malignancies, where it is likely to play a role in the malignant phenotype: undifferentiated nasopharyngeal carcinoma (100% of cases) and gastric carcinomas (about 10% of cases). We and others have obtained growth transformation of monkey kidney primary epithelial cells by transfection of viral DNA, especially with the BARF1 gene of EBV (Wei et al., 1997). We now report that the same type of primary epithelial cells can be growth-transformed using EBV particles derived from a nasopharyngeal carcinoma tumor line. Not only can these EBV-infected cells grow over 100 passages, escaping senescence, in contrast to their noninfected counterparts, but they can also survive and proliferate at very low cell density. Several subclones were characterized in terms of viral gene expression. All these clones gave a similar pattern, with detection of EBNA1 and BARF1 proteins but absence of LMP1. CD21, which is the main EBV receptor on B lymphocytes, was not expressed on parental monkey kidney epithelial cells nor on EBV-infected cell clones. This model of epithelial cell transformation will be useful for a better investigation of EBV functions critical for oncogenesis of epithelial cells.

[The biology of nasopharyngeal carcinoma in 2001: update and perspective]. / La biologie des carcinomes nasopharyngés en 2001: mise à jour et perspectives.

The phenotype of malignant epithelial cells in nasopharyngeal carcinomas (NPC) results from latent infection by the Epstein-Barr virus (EBV) combined to cell gene alterations, especially those affecting the p16/Ink 4 gene. At the site of the primary tumor, NPC are strongly infiltrated by non-malignant EBV-negative T-lymphocytes. There are experimental clues suggesting that these lymphocytes are involved in tumor development, for example by providing anti-apoptotic signals to malignant epithelial cells. The amazing geographic distribution of NPC is accounted for by the conjunction of several risk factors in endemic regions. These risk factors are related to the diffusion of one or several susceptibility genes, the probable existence of viral strains with high oncogenic potential and non-viral environmental factors, especially dietary factors. The perspectives of immunotherapy in NPC are still unclear since viral proteins detected in tumors are poorly immunogenic (EBNA1, LMP1). Targeted molecules designed to interfere with viral and cellular oncoprotein signals will probably have interesting applications for the treatment of NPC.

Contrasted frequencies of p53 accumulation in the two age groups of North African nasopharyngeal carcinomas.

EBV-associated nasopharyngeal carcinomas (NPCs) from Southeast Asia and North Africa have many common clinical and biological characteristics. However, they differ with regard to their age distribution. In Asia, NPC mainly affects patients in the 4th or 5th decade of their life, whereas in North Africa an additional peak of incidence is found between the ages of 10 and 20. The p53 gene is rarely mutated in NPC. However, several groups have reported a consistent accumulation of p53 in Asian NPCs. To determine whether p53 was also accumulated in North African NPCs, we investigated its expression, by immunohistochemistry, in a series of 90 Tunisian biopsies. Bc12 and CD95, two proteins involved in the regulation of cell survival and apoptosis, were investigated in the same study. We found accumulation of p53 in 81% of the cases for patients over 30 years of age, but in only 38% of specimens for younger patients (P = 0.00013). There was a trend toward a higher frequency of Bc12 detection in patients over 30, but it was not statistically significant. CD95 expression was detected in all biopsies, generally at a high level, even at advanced stages of the disease. The changing frequency of p53 accumulation, below and over 30, suggests that NPC cells often achieve malignant transformation through different pathways in both age groups.

Radiation-induced expression of functional Fas ligand in EBV-positive human nasopharyngeal carcinoma cells.

Ionizing radiation remains a major therapeutic tool against human cancers, especially epithelial tumors, which account for the majority of human malignancies. Although Fas and Fas-L are essential determinants of apoptosis, few data support their role in the cytotoxic effect of ionizing radiation. Epstein-Barr-virus (EBV)-positive nasopharyngeal carcinoma (NPC) were chosen to address this question owing to their known sensitivity to ionizing radiation and their constitutive expression of the Fas-receptor. We here report that, in xenografted NPC cells, Fas-L expression, which was very low in basal conditions, was dramatically increased by tumor irradiation. Both the Fas receptor and the Fas ligand were found to be functional in this model, and a high proportion of irradiated NPC cells underwent apoptosis following tumor irradiation. Induction of Fas-L expression and apoptosis were observed for doses as low as 2 Gy. These data show an increase in Fas-L expression upon irradiation exposure, and strongly suggest that, in some epithelial malignancies, Fas-mediated apoptosis can play a major role in the anti-tumor effect of ionizing radiation, in the range of doses used for therapeutic applications.

Nitrous oxide anesthesia-associated myelopathy.

BACKGROUND: The role of nitrous oxide exposure in neurologic complications of subclinical cobalamin deficiency has been reported, but few cases are well documented. OBSERVATION: Two weeks after surgery for prosthetic adenoma, a 69-year-old man developed ascending paresthesia of the limbs, severe ataxia of gait, tactile sensory loss on the 4 limbs and trunk, and absent tendon reflexes. After a second surgical intervention, the patient became confused. Four months after onset, the patient had paraplegia, severe weakness of the upper limbs, cutaneous anesthesia sparing the head, and confusion. Moderate macrocytosis, low serum B12 levels, and a positive Schilling test result led to the diagnosis of pernicious anemia. Results of electrophysiologic examinations showed a diffuse demyelinating neuropathy. Magnetic resonance imaging of the spinal cord disclosed hyperintensities of the dorsal columns on T2-weighted images. CONCLUSIONS: Pernicious anemia can result in severe neurologic symptoms with only mild hematologic changes. The role of nitrous oxide anesthesia in revealing subclinical B12 deficiency must be emphazised. Magnetic resonance imaging of the spinal cord might be helpful in making the diagnosis.

Evidence of LMP1-TRAF3 interactions in glycosphingolipid-rich complexes of lymphoblastoid and nasopharyngeal carcinoma cells.

Latent membrane protein 1 (LMP1) is an Epstein-Barr virus (EBV) protein expressed in EBV-transformed B lymphocytes and in approximately 50% of nasopharyngeal carcinomas (NPCs). LMP1 signaling involves several cellular signaling intermediates, especially TNF receptor-associated factors (TRAFs). We have shown previously that LMP1 is highly concentrated in a cell fraction called glycosphingolipid-rich membrane complexes (GSL complexes). We report here that parallel accumulation of LMP1 and TRAF3, but not TRAF1 or TRADD, was observed in GSL complexes from lymphoblastoid and LMP1-positive NPC cells. In contrast, TRAF3 was not concentrated in GSL complexes from LMP1-negative cells. Binding of LMP1 and TRAF3 in GSL complexes was demonstrated in lymphoblastoid and NPC cells, by co-immunoprecipitation with both anti-LMP1 and anti-TRAF3 antibodies.

Control of apoptosis in Epstein Barr virus-positive nasopharyngeal carcinoma cells: opposite effects of CD95 and CD40 stimulation.

The expression and function of CD95 and CD40 were investigated in malignant cells from EBV-positive undifferentiated nasopharyngeal carcinomas (NPCs). Large amounts of CD95 and CD40 expression were detected in 15 of 16 EBV-positive NPC specimens. In contrast, CD95 was not detected in two biopsies from patients with EBV-negative differentiated NPCs. We tested whether the CD95 apoptotic pathway was functional in NPC cells by treating two EBV-positive NPC tumor lines in vitro with a CD95 agonist. In both cases, NPC cells were extremely susceptible to CD95-mediated apoptosis, despite strong constitutive expression of Bcl-x. Combined CD40 and CD95 stimulation was used to investigate the possible anti-apoptotic activity mediated by CD40. The CD40 receptor was activated by incubating NPC cells with murine L cells producing CD154, the CD40 ligand. This treatment resulted in a strong inhibition of CD95-related cytotoxicity. Such an anti-apoptotic effect of CD40 is well known for B lymphocytes, but has not previously been reported for epithelial cells. These data suggest that NPC tumor-infiltrating lymphocytes, which often produce the CD40 ligand in situ, may increase the survival of malignant cells, thereby enhancing tumor growth in patients.

Molecular characterization of antigen-processing function in nasopharyngeal carcinoma (NPC): evidence for efficient presentation of Epstein-Barr virus cytotoxic T-cell epitopes by NPC cells.

Potentiation of the EBV-specific CTL response by immunization with CTL epitopes has been proposed as a logical approach for immune-targeting nasopharyngeal carcinoma (NPC) cells in vivo. This approach will undoubtedly be influenced by the ability of these malignant cells to endogenously process and present target epitopes on their cell surface for immune recognition by CTLs. Analysis of NPC cells in fresh tumor biopsies and long-term, established NPC tumors in nude mice revealed normal expression of the MHC-encoded putative peptide transporters TAP1 and TAP2, as well as the proteasome components LMP2 and LMP7, which have been shown previously to be essential components of the class I processing pathway. Moreover, these tumor cells also showed high levels of HLA class I alleles on the cell surface, suggesting that peptides are available for binding to nascent MHC molecules in the endoplasmic reticulum. Using a recombinant vaccinia virus to transiently express the EBV nuclear antigens, we studied the antigen-processing efficiency of NPC cells. Our findings demonstrate that, in contrast to cells from another EBV-associated malignancy, Burkitt's lymphoma, NPC cells display normal antigen-processing function and are efficiently recognized by HLA class I-restricted, virus-specific CTLs. These studies also provide a rationale for focusing on strategies designed to activate CTLs specific for EBV antigens that are expressed in NPC cells in vivo.

[Multiple and unilateral paralysis of the cranial nerves revealing 2 cerebellar infarctions]. / Paralysies multiples et unilatérales des nerfs crâniens révélant deux infarctus cérébelleux.

A 86 year old man suffered multiple palsies of the right V, VI, VII, VIII, IX and X cranial nerves preceded for several weeks by transient diplopia, facial palsy and vertigo. The CT scan and MRI showed two infarcts sitting in the territories of the right postero-inferior cerebellar artery and the right antero-inferior cerebellar artery. A cerebellar syndrome developed several days later. Although cranial nerves palsies are very commun following infarcts of the cerebellar arteries, their occurrence without other neurological deficit, especially cerebellar syndrome, seems to be rare.

High concentration of the EBV latent membrane protein 1 in glycosphingolipid-rich complexes from both epithelial and lymphoid cells.

Latent Membrane Protein 1 (LMP1) is an EBV-transforming protein which is detected both in lymphoblastoid cell lines-resulting from EBV-immortalization in vitro- and in undifferentiated nasopharyngeal carcinoma (NPC), an EBV-associated malignancy of epithelial origin. To better define LMP1 subcellular targets, LMP1 distribution was analyzed in cellular glycosphingolipid-rich complexes (GSL-complexes) derived from epithelial and lymphoid cells. These complexes are obtained by extraction of glycosphingolipid-rich membrane domains (GSL-domains), which are clustering sites for heterotrimeric G-proteins and G-protein-associated receptors. LMP1 concentration was enriched 50-fold in GSL-complexes extracted from a NPC tumor line, C15. High concentrations of LMP1 were also observed in GSL-complexes derived from cultured lymphoid and epithelial cells. These data suggest that association with GSL-domains is an important step in LMP1 trafficking and is probably required for some aspects of its biological activity.

Chromosome 11q13 gene amplifications in oral and oropharyngeal carcinomas: no correlation with subclinical lymph node invasion and disease recurrence.

Gene amplifications in the q13 band of chromosome 11 are among the most frequent genetic alterations in head and neck squamous cell carcinomas. Previous studies have suggested that such amplification is a marker of aggressive tumor evolution. Their potential for predicting subclinical lymph node invasion or disease recurrence was investigated in a prospective series of 50 oral and oropharyngeal carcinomas. Cell DNA content was also measured in 32 tumors of this series. Gene amplifications affecting the 11q13 band were detected in 11 of 50 (20%) patients, a relatively low frequency in comparison with data reported previously for other carcinomas of the upper aerodigestive tract, especially hypopharyngeal carcinomas. These gene amplifications were preferentially associated with aneuploidy. Cervical lymph nodes of 26 clinically N0 (Tumor-Node-Metastasis staging) patients were surgically explored. The frequency of 11q13 amplifications was very similar in the presence or in the absence of histological invasion, 3 of 15 (20%) and 2 of 11 (18%), respectively. Thus, 11q13 amplifications do not appear to be a reliable marker for prediction of subclinical lymph-node invasion in oral and oropharyngeal carcinomas. The detection of 11q13 amplifications was also not associated with a higher risk of disease recurrence. These data suggest that not only the prevalence but also the prognostic significance of 11q13 amplifications varies between tumors at different sites in the upper aerodigestive tract.